Noncompetitive immunoassay detection system for haptens on the basis of antimetatype antibodies.
Clin Chem
; 61(4): 627-35, 2015 Apr.
Article
en En
| MEDLINE
| ID: mdl-25695852
ABSTRACT
BACKGROUND:
Small molecules classified as haptens are generally measured by competitive immunoassay, which is theoretically inferior to noncompetitive sandwich immunoassay in terms of sensitivity and specificity. We created a method for developing sandwich immunoassays to measure haptens on the basis of antimetatype antibodies.METHODS:
We generated antimetatype monoclonal antibodies against a hapten-antibody immunocomplex using an ex vivo antibody development system, the Autonomously Diversifying Library (ADLib) system. We selected 2 haptens, estradiol (E2) and 25-hydroxyvitamin D [25(OH)D], as analytes. Sandwich immunoassays for these 2 haptens were developed by use of a 96-well microtiter plate and a fully automated chemiluminescence analyzer, and the performances of these immunoassays were investigated.RESULTS:
The developed assays exhibited sensitivity high enough to detect target haptens in serum samples. The limit of detection of the ELISA for E2 was 3.13 pg/mL, and that of the fully automated chemiluminescent enzyme immunoassay (CLEIA) system was 2.1 ng/mL for 25(OH)D. The cross-reactivity with immunoreactive derivatives was effectively improved compared with the competitive assay. The CVs for the sandwich ELISA for E2 were 4.2%-12.6% (intraassay) and 6.2%-21.8% (total imprecision). The CVs for the sandwich CLEIA for 25(OH)D were 1.0%-2.3% (intraassay) and 1.9%-3.5% (total imprecision). In particular, the sandwich CLEIA for 25(OH)D showed correlations of r = 0.99 with both LC-MS/MS and a commercially available (125)I RIA.CONCLUSIONS:
Our method represents a potentially simple and practical approach for routine assays of haptens, including vitamins, hormones, drugs, and toxins.
Texto completo:
1
Base de datos:
MEDLINE
Asunto principal:
Vitamina D
/
Inmunoensayo
/
Estradiol
/
Haptenos
/
Anticuerpos Monoclonales
Tipo de estudio:
Diagnostic_studies
Idioma:
En
Revista:
Clin Chem
Asunto de la revista:
QUIMICA CLINICA
Año:
2015
Tipo del documento:
Article